Sexual function and pelvic floor function in men with systemic sclerosis compared to healthy controls: a cross-sectional study.

OBJECTIVES: This cross-sectional study aimed to compare the sexual function (SF) and pelvic floor function of men with systemic sclerosis (SSc) with age-matched healthy controls (HC) and to identify the implications of clinical features on SF. MATERIAL AND METHOD: Twenty SSc males and 20 HC aged 18-70 years completed eleven questionnaires assessing SF [International Index of Erectile Function (IIEF), Male Sexual Health Questionnaire (MSHQ)]; sexual quality of life: Sexual Quality of Life Questionnaire-Male (SQoL-M); pelvic floor function: Pelvic Floor Impact Questionnaire-Short Form 7 (PFIQ-7), fatigue, depression, physical fitness, functional disability, and quality of life. Clinical data were collected. RESULTS: Significantly worse SF was observed in patients (median IIEF erectile function 12 in SSc versus 29 in HC, p < 0.001), with 70% reporting erectile dysfunction (ED) compared to 15% in HC. However, no significant difference was observed regarding pelvic floor function (median PFIQ7 8.8 in SSc versus 7.0 in HC, p = 0.141). Impaired SF was associated with higher disease activity, increased systemic inflammation, more pronounced fatigue, reduced physical fitness, severe depression, impaired overall quality of life, dyspepsia, and arthralgias (p < 0.05 for all). CONCLUSIONS: Sexual dysfunction is highly prevalent in our SSc patients, whereas pelvic floor dysfunction is unlikely to be associated with these problems.

Biomarker analysis from the phase 2b randomized placebo-controlled trial of riociguat in early diffuse cutaneous systemic sclerosis.

OBJECTIVE: To examine disease and target engagement biomarkers in the RISE-SSc trial of riociguat in early diffuse cutaneous systemic sclerosis and their potential to predict the response to treatment. METHODS: Patients were randomized to riociguat (n = 60) or placebo (n = 61) for 52 weeks. Skin biopsies and plasma/serum samples were obtained at baseline and week 14. Plasma cyclic guanosine monophosphate (cGMP) was assessed using radio-immunoassay. Alpha smooth muscle actin (αSMA) and skin thickness were determined by immunohistochemistry, mRNA markers of fibrosis by qRT-PCR in skin biopsies, and serum CXC motif chemokine ligand 4 (CXCL-4) and soluble platelet endothelial cell adhesion molecule-1 (sPECAM-1) by enzyme-linked immunosorbent assay. RESULTS: By week 14, cGMP increased by 94 ± 78% with riociguat and 10 ± 39% with placebo (p < 0.001, riociguat vs placebo). Serum sPECAM-1 and CXCL-4 decreased with riociguat vs placebo (p = 0.004 and p = 0.008, respectively). There were no differences in skin collagen markers between the 2 groups. Higher baseline serum sPECAM-1 or the detection of αSMA-positive cells in baseline skin biopsies were associated with a larger reduction of modified Rodnan skin score from baseline at week 52 with riociguat vs placebo (interaction P-values 0.004 and 0.02, respectively). CONCLUSION: Plasma cGMP increased with riociguat, suggesting engagement with the nitric oxide-soluble guanylate cyclase-cGMP pathway. Riociguat was associated with a significant reduction in sPECAM-1 (an angiogenic biomarker) vs placebo. Elevated sPECAM-1 and the presence of αSMA-positive skin cells may help to identify patients who could benefit from riociguat in terms of skin fibrosis. TRIAL REGISTRATION: Clinicaltrials.gov, NCT02283762.

S100A4-neutralizing monoclonal antibody 6B12 counteracts the established experimental skin fibrosis induced by bleomycin.

OBJECTIVES: Our previous studies have demonstrated that the Damage Associated Molecular Pattern (DAMP) protein, S100A4, is overexpressed in the involved skin and peripheral blood of patients with SSc. It is associated with skin and lung involvement, and disease activity. By contrast, lack of S100A4 prevented the development of experimental dermal fibrosis. Herein we aimed to evaluate the effect of murine anti-S100A4 mAb 6B12 in the treatment of preestablished experimental dermal fibrosis. METHODS: The effects of 6B12 were assessed at therapeutic dosages in a modified bleomycin-induced dermal fibrosis mouse model by evaluating fibrotic (dermal thickness, proliferation of myofibroblasts, hydroxyproline content, phosphorylated Smad3-positive cell count) and inflammatory (leukocytes infiltrating the lesional skin, systemic levels of selected cytokines and chemokines) outcomes, and transcriptional profiling (RNA sequencing). RESULTS: Treatment with 7.5 mg/kg 6B12 attenuated and might even reduce pre-existing dermal fibrosis induced by bleomycin as evidenced by reduction in dermal thickness, myofibroblast count and collagen content. These antifibrotic effects were mediated by the downregulation of TGF-ß/Smad signalling and partially by reducing the number of leukocytes infiltrating the lesional skin and decrease in the systemic levels of IL-1α, eotaxin, CCL2 and CCL5. Moreover, transcriptional profiling demonstrated that 7.5 mg/kg 6B12 also modulated several profibrotic and proinflammatory processes relevant to the pathogenesis of SSc. CONCLUSION: Targeting S100A4 by the 6B12 mAb demonstrated potent antifibrotic and anti-inflammatory effects on bleomycin-induced dermal fibrosis and provided further evidence for the vital role of S100A4 in the pathophysiology of SSc.

Does the Impact of COVID-19 on Patients With Systemic Sclerosis Change Over Time?

OBJECTIVE: The outcome of patients with COVID-19 improved over the pandemic, including patients with systemic rheumatic diseases. However, data on patients with systemic sclerosis (SSc) are lacking. This study aimed to assess the outcome of patients with both SSc and COVID-19 over several waves. METHODS: Patients with both SSc and COVID-19 who were registered in the European Scleroderma Trials and Research group (EUSTAR) were collected between April 2020 and April 2021. Patients were assigned to waves 1, 2, or 3 depending on the date of their COVID-19 diagnosis. Primary endpoints were death, intensive care unit stay, or ventilatory support (severe outcome). Subgroup analyses of patients who were hospitalized or died were conducted. General and SSc-specific characteristics and treatment were compared over the waves. Descriptive statistics and multivariate logistic regression were applied. RESULTS: A total of 333 patients were included; 57 patients (17%) had a severe outcome, and 30 patients (9%) died. Compared to wave 1, significantly fewer patients with SSc suffered from severe COVID-19 in waves 2 and 3 (28.2% vs 9.8% and 12.7%; P < 0.001), fewer patients required hospitalization (46.7% vs 19.6% and 25.5%; P < 0.001) or ventilatory support (24.0% vs 8.7% and 10.9%; P = 0.001), and fewer patients died (15.7% vs 5.0% and 7.5%; P = 0.011). Patients were significantly younger, more often men, had less frequent arterial hypertension, and less SSc cardiac involvement over waves 1 to 3. Patients received significantly less medium to high doses of corticosteroids as they did SSc treatment. CONCLUSION: The outcome of patients with both SSc and COVID-19 improved significantly over time because of intrinsic and extrinsic factors.

Correction: Storkánová et al. Inhibition of Hsp90 Counteracts the Established Experimental Dermal Fibrosis Induced by Bleomycin. Biomedicines 2021, 9, 650.

In the original publication [...].

Effect of an 8-Week Tailored Physiotherapy Program on Sexual Health in Women with Scleroderma and Myositis: A Controlled Pilot Study.

INTRODUCTION: Systemic sclerosis (SSc) and idiopathic inflammatory myopathies (IIM) are very rare rheumatic diseases burdened by a high prevalence of sexual dysfunctions. However, no specific treatment has been proposed to date. To our knowledge, this is the first (pilot) study aiming to investigate the effect of an 8-week tailored physiotherapy program on the sexual health of women with SSc and IIM. METHODS: In total, 12 women with SSc and 4 women with IIM were enrolled in the study. Based on the patients' capability to participate in the program, they were divided into an intervention group (IG) (mean ± SD age 46.8 ± 8.6 years) and a control group (CG) (mean ± SD age 46.3 ± 8.5 years). IG underwent the 8-week program (1 h of supervised physiotherapy twice weekly), whereas CG received no physiotherapy. At weeks 0 and 8, all patients filled in questionnaires assessing sexual function (Female Sexual Function Index [FSFI], Brief Index of Sexual Functioning for Women [BISF-W]), sexual quality of life (Sexual Quality of Life-Female [SQoL-F]), functional ability (Health Assessment Questionnaire [HAQ]), quality of life (Medical Outcomes Short Form-36 [SF-36]), and depression (Beck's Depression Inventory-II [BDI-II]). The changes were analyzed with two-way ANOVA and Friedmann's test. RESULTS: Compared to the statistically significant deterioration in CG over weeks 0-8, we found statistically significant improvements in the total scores of FSFI and BISF-W, and some of their domains, functional status, and the physical component of quality of life. CONCLUSION: Our 8-week physiotherapy program not only prevented the natural course of progressive deterioration of functional ability but also led to a significant improvement in sexual function and quality of life in women with SSc and IIM. However, due to the lack of randomization and a relatively small sample size resulting from the strict inclusion criteria, further validation of our results is needed. TRIAL REGISTRATION NUMBER: ISRCTN91200867 (prospectively registered).

Riociguat in patients with early diffuse cutaneous systemic sclerosis (RISE-SSc): open-label, long-term extension of a phase 2b, randomised, placebo-controlled trial.

BACKGROUND: The phase 2b Riociguat Safety and Efficacy in Patients with Diffuse Cutaneous Systemic Sclerosis (RISE-SSc) trial investigated riociguat versus placebo in early diffuse cutaneous systemic sclerosis. The long-term extension evaluated safety and exploratory treatment effects for an additional year. METHODS: Patients were enrolled to RISE-SSc between Jan 15, 2015, and Dec 8, 2016. Those who completed the 52-week, randomised, parallel-group, placebo-controlled, double-blind phase were eligible for the long-term extension. Patients originally assigned to riociguat continued therapy (riociguat-riociguat group). Those originally assigned to placebo were switched to riociguat (placebo-riociguat group), adjusted up to 2·5 mg three times daily in a 10-week, double-blind dose-adjustment phase, followed by an open-label phase. Statistical analyses were descriptive. Safety including adverse events and serious adverse events was assessed in the long-term safety analysis set (all patients randomly assigned and treated with study medication in the double-blind phase who continued study medication in the long-term extension). The RISE-SSc trial is registered with ClinicalTrials.gov, NCT02283762. FINDINGS: In total, 87 (72%) of 121 patients in the main RISE-SSc study entered the long-term extension (riociguat-riociguat, n=42; placebo-riociguat, n=45). 65 (75%) of 87 patients were women, 22 (25%) were men, and 62 (71%) were White. Overall, 82 (94%) of 87 patients in the long-term extension had an adverse event; most (66 [76%] of 87) were of mild to moderate severity, with no increase in pulmonary-related serious adverse events in patients with interstitial lung disease. INTERPRETATION: No new safety signals were observed with long-term riociguat in patients with early diffuse cutaneous systemic sclerosis. Study limitations include the absence of a comparator group in this open-label extension study. FUNDING: Bayer and Merck Sharp & Dohme.

The effect of a 24-week physiotherapy and occupational therapy program in systemic sclerosis: a monocentric controlled study with follow-up.

OBJECTIVES: The structural and functional changes of the hands and face in systemic sclerosis (SSc) can be severely disabling. We aimed to assess the effect of a 24-week supervised physiotherapy and occupational therapy program (POTp) combined with home exercise on the function of hands/mouth of SSc patients, compared to a daily home exercise program in typical outpatient care. METHODS: Fifty-nine patients with SSc were consecutively and non-selectively enrolled in an intervention (IG, n=27) or control (CG, n=32) group. Only the IG underwent the POTp twice a week for 1.5 hours. At baseline, 12, 24, and 48 weeks, all patients were assessed by a blinded physiotherapist for the hands/mouth function (delta finger-to-palm, handgrip strength, Hand and Mobility in Scleroderma, interincisal/interlabial distance), and self-evaluated their hand (Cochin Hand Function Scale) and mouth function (Mouth Handicap in Systemic Sclerosis scale), disability (Health Assessment Questionnaire [HAQ], SSc HAQ), and quality of life (Short Form-36). RESULTS: At week 24, compared to the significant deterioration in the CG, we found a significant improvement in the IG in the objectively assessed hands/mouth function and in the subjectively evaluated hand function and disability. The improvement was clinically meaningful (by >20%) in a substantial proportion of patients. Although the improvement in most outcomes was still present at week 48, the maximum effect was not sustained. CONCLUSIONS: This 24-week POTp not only attenuated the progressive deterioration, but also significantly improved the function of the hands/mouth, which was clinically meaningful in a substantial proportion of patients with SSc.

Female Sexual Dysfunction and Pelvic Floor Muscle Function Associated with Systemic Sclerosis: A Cross-Sectional Study.

Only a few studies have addressed sexual health in patients with systemic sclerosis (SSc). This study aimed to compare female sexual function and pelvic floor muscle function in SSc patients with healthy controls (HC) matched by age, and to identify the potential implications of clinical features on sexual function. Our cohort included 90 women with SSc and 90 HC aged 18-70 years that completed six well-established and validated questionnaires assessing sexual function (Brief Index of Sexual Function for Women, Female Sexual Function Index, Sexual Quality of Life Questionnaire-Female, Sexual Function Questionnaire) and pelvic floor function (Pelvic Floor Impact Questionnaire-Short Form 7 and Pelvic Organ Prolapse/Urinary Incontinence Sexual Questionnaire Short Form). Results from women with SSc and HC were contrasted and correlated with relevant clinical features. The prevalence of female sexual dysfunction was 73% in SSc patients (vs. 31% in HC). Women with SSc reported significantly worse pelvic floor function and sexual function than HC. Impaired sexual function was correlated with higher disease activity, the presence of dyspnea and interstitial lung disease, increased systemic inflammation, reduced physical activity, functional disability, more severe depression, more pronounced fatigue, and impaired quality of life. We demonstrate that sexual dysfunction is highly prevalent among women with SSc. This aspect of the disease deserves more attention both in clinical care and at the level of international research collaboration.

Inhibition of Hsp90 Counteracts the Established Experimental Dermal Fibrosis Induced by Bleomycin.

Our previous study demonstrated that heat shock protein 90 (Hsp90) is overexpressed in the involved skin of patients with systemic sclerosis (SSc) and in experimental dermal fibrosis. Pharmacological inhibition of Hsp90 prevented the stimulatory effects of transforming growth factor-beta on collagen synthesis and the development of dermal fibrosis in three preclinical models of SSc. In the next step of the preclinical analysis, herein, we aimed to evaluate the efficacy of an Hsp90 inhibitor, 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), in the treatment of established experimental dermal fibrosis induced by bleomycin. Treatment with 17-DMAG demonstrated potent antifibrotic and anti-inflammatory properties: it decreased dermal thickening, collagen content, myofibroblast count, expression of transforming growth factor beta receptors, and pSmad3-positive cell counts, as well as leukocyte infiltration and systemic levels of crucial cytokines/chemokines involved in the pathogenesis of SSc, compared to vehicle-treated mice. 17-DMAG effectively prevented further progression and may induce regression of established bleomycin-induced dermal fibrosis to an extent comparable to nintedanib. These findings provide further evidence of the vital role of Hsp90 in the pathophysiology of SSc and characterize it as a potential target for the treatment of fibrosis with translational implications due to the availability of several Hsp90 inhibitors in clinical trials for other indications.

Plasma Hsp90 levels in patients with systemic sclerosis and relation to lung and skin involvement: a cross-sectional and longitudinal study.

Our previous study demonstrated increased expression of Heat shock protein (Hsp) 90 in the skin of patients with systemic sclerosis (SSc). We aimed to evaluate plasma Hsp90 in SSc and characterize its association with SSc-related features. Ninety-two SSc patients and 92 age-/sex-matched healthy controls were recruited for the cross-sectional analysis. The longitudinal analysis comprised 30 patients with SSc associated interstitial lung disease (ILD) routinely treated with cyclophosphamide. Hsp90 was increased in SSc compared to healthy controls. Hsp90 correlated positively with C-reactive protein and negatively with pulmonary function tests: forced vital capacity and diffusing capacity for carbon monoxide (DLCO). In patients with diffuse cutaneous (dc) SSc, Hsp90 positively correlated with the modified Rodnan skin score. In SSc-ILD patients treated with cyclophosphamide, no differences in Hsp90 were found between baseline and after 1, 6, or 12 months of therapy. However, baseline Hsp90 predicts the 12-month change in DLCO. This study shows that Hsp90 plasma levels are increased in SSc patients compared to age-/sex-matched healthy controls. Elevated Hsp90 in SSc is associated with increased inflammatory activity, worse lung functions, and in dcSSc, with the extent of skin involvement. Baseline plasma Hsp90 predicts the 12-month change in DLCO in SSc-ILD patients treated with cyclophosphamide.

Primary vasculitides - current diagnostics and therapy.

By term „vasculitis“ inflammatory disease of blood vessels is designated that leads to vessel wall destruction followed by proliferation and occlusion of their lumina. Basic condition for this diagnosis is that vessel wall is a primary site of the pathological process. Clinical syndromes are a consequence of this process resulting into ischaemia of tissues supplied by the affected vessels and with constitutional symptoms associated to the inflammatory disease. Vasculitis can occur de novo as a primary involvement of vessel wall of unknown aetiology or it develops secondary to other diseases. The diagnosis of vasculitis is usually based on pathological findings from biopsy or abnormalities detected by imaging methods. This review describes the common primary vasculitides, their diagnostics and management.

Secondary vasculitis - omitted manifestation of many diseases.

Secondary vasculitides usually accompany various common and rare conditions, Their clinical picture is very diverse, they can be loclaized or genaralized. Most frequently, we find parainfectious, drug-related and paraneoplastic vasculitides, less commonly in connective tissue diseases, after radiotherapy or transplantation. Vasculitides may be associated to infection of any origin. Drug-related vasculitides are mainly confined to the skin with picture of leukocytoclastic angiitis but visceral organs may be involved too. Paraneoplastic vasculitides usually accompany solid tumours and lymphoproliferative processes. When related to connective tissue diseases we can observe vasculitis in rheumatoid arthritis, systemic lupus erythematosus, Sjoegren syndrome, systemic sclerosis and other conditions. The diagnosis of vasculitis is usually based on pathological findings from biopsy. Management lies in treatment of underlying disease and if it is ineffective combined immunosuppression should be introduced.

Riociguat in patients with early diffuse cutaneous systemic sclerosis (RISE-SSc): randomised, double-blind, placebo-controlled multicentre trial.

OBJECTIVES: Riociguat is approved for pulmonary arterial hypertension and has antiproliferative, anti-inflammatory and antifibrotic effects in animal models of tissue fibrosis. We evaluated the efficacy and safety of riociguat in patients with early diffuse cutaneous systemic sclerosis (dcSSc) at high risk of skin fibrosis progression. METHODS: In this randomised, double-blind, placebo-controlled, phase IIb trial, adults with dcSSc of <18 months' duration and a modified Rodnan skin score (mRSS) 10-22 units received riociguat 0.5 mg to 2.5 mg orally three times daily (n=60) or placebo (n=61). The primary endpoint was change in mRSS from baseline to week 52. RESULTS: At week 52, change from baseline in mRSS units was -2.09±5.66 (n=57) with riociguat and -0.77±8.24 (n=52) with placebo (difference of least squares means -2.34 (95% CI -4.99 to 0.30; p=0.08)). In patients with interstitial lung disease, forced vital capacity declined by 2.7% with riociguat and 7.6% with placebo. At week 14, average Raynaud's condition score had improved ≥50% in 19 (41.3%)/46 patients with riociguat and 13 (26.0%)/50 patients with placebo. Safety assessments showed no new signals with riociguat and no treatment-related deaths. CONCLUSIONS: Riociguat did not significantly benefit mRSS versus placebo at the predefined p<0.05. Secondary and exploratory analyses showed potential efficacy signals that should be tested in further trials. Riociguat was well tolerated.

[Novel trends in monitoring and therapy of ANCA associated vasculitides]. / Nové trendy v monitorování a lécbe ANCA asociovaných vaskulitid.

Vasculitides with positivity of autoantibodies to neutrophil leukocytes cytoplasm (ANCA, AAV) belong to primary vasculitides involving small and less commonly medium size blood vessels. Three different clinical types of AAV can be distinguished: granulomatosis with polyangiitis, eosinophilic granulomatosis with polyangiitis and microscopic polyangiitis. Since these autoantibodies seem to be weak activity biomarkers of AAV new molecules and factors start to come up, e.g. neutrophil extracellular traps NET, several T-lymphocyte subpopulations and different immunoglobulins classes of ANCA. In modern biological therapy rituximab is widely used, for refractory cases intravenous immunoglobulins and antithymocyte globulin are recommended. The data from clinical trials with alemtuzumab are controversial, but avacopan selective inhibitor of C5a receptor and inhibitor of B-lymphocyte activation factor belimumab are promise for future.Key words: biologicals - biomarkers - eosinophilic granulomatosis with polyangiitis - granulomatosis with polyangiitis - microscopic polyangiitis.

Mapping and predicting mortality from systemic sclerosis.

OBJECTIVES: To determine the causes of death and risk factors in systemic sclerosis (SSc). METHODS: Between 2000 and 2011, we examined the death certificates of all French patients with SSc to determine causes of death. Then we examined causes of death and developed a score associated with all-cause mortality from the international European Scleroderma Trials and Research (EUSTAR) database. Candidate prognostic factors were tested by Cox proportional hazards regression model by single variable analysis, followed by a multiple variable model stratified by centres. The bootstrapping technique was used for internal validation. RESULTS: We identified 2719 French certificates of deaths related to SSc, mainly from cardiac (31%) and respiratory (18%) causes, and an increase in SSc-specific mortality over time. Over a median follow-up of 2.3 years, 1072 (9.6%) of 11 193 patients from the EUSTAR sample died, from cardiac disease in 27% and respiratory causes in 17%. By multiple variable analysis, a risk score was developed, which accurately predicted the 3-year mortality, with an area under the curve of 0.82. The 3-year survival of patients in the upper quartile was 53%, in contrast with 98% in the first quartile. CONCLUSION: Combining two complementary and detailed databases enabled the collection of an unprecedented 3700 deaths, revealing the major contribution of the cardiopulmonary system to SSc mortality. We also developed a robust score to risk-stratify these patients and estimate their 3-year survival. With the emergence of new therapies, these important observations should help caregivers plan and refine the monitoring and management to prolong these patients' survival.

Molecular markers of systemic autoimmune disorders: the expression of MHC-located HSP70 genes is significantly associated with autoimmunity development.

OBJECTIVES: To analyse the expression regulation of two inducible HSP70 genes - HSPA1A and HSPA1B - located within the major histocompatibility complex (MHC) in patients with various systemic autoimmune diseases and to prove the reliability of MHC-located HSP70 genes as molecular markers reflecting the autoimmune process. METHODS: 94 adult patients with idiopathic inflammatory myopathy (IIM, n=31), systemic lupus erythematosus (SLE, n=31) or systemic sclerosis (SSc, n=32) and 37 healthy individuals were analysed. The mRNA expression level was determined using quantitative real-time PCR method. The expression of intracellular HSP70 was established by flow cytometry, the extracellular HSP70 protein was measured in plasma samples using a commercially available sandwich enzyme-linked immunosorbent assay (ELISA). RESULTS: The expression of HSPA1A gene was significantly up-regulated in patients with autoimmune diseases (SLE: p<0.01; SSc: p<0.01; IIM: p<0.0001) compared to healthy controls. The expression of HSPA1B gene was increased only in patients with myositis (p<0.05). Furthermore, the HSPA1B gene expression is associated with the HLA-DRB1*03 risk allele in patients with IIM. In addition, we have found a relation between HSPA1A gene expression regulation and the presence of disease specific autoantibodies in patients with SLE and myositis. The level of intracellular HSP70 was not increased; however, the level of extracellular HSP70 protein was increased in patients suffering from SSc and IIM as compared to controls. CONCLUSIONS: The results suggest an involvement of the MHC-linked HSP70 genes in the pathology of studied autoimmune disorders. Therefore, the HSPA1A and HSPA1B genes might serve as an interesting candidate molecule for development of distinct types of autoimmunities.

Tribbles homologue 3 stimulates canonical TGF-ß signalling to regulate fibroblast activation and tissue fibrosis.

OBJECTIVES: Tribbles homologue 3 (TRB3) is a pseudokinase that modifies the activation of various intracellular signalling pathways to control fundamental processes extending from mitosis and cell activation to apoptosis and modulation of gene expression. Here, we aimed to analyse the role of TRB3 in fibroblast activation in systemic sclerosis (SSc). METHODS: The expression of TRB3 was quantified by quantitative PCR, western blot and immunohistochemistry. The role of TRB3 was analysed in cultured fibroblasts and in experimental fibrosis using small interfering RNA (siRNA)-mediated knockdown and overexpression of TRB3. RESULTS: TRB3 expression was increased in fibroblasts of patients with SSc and in murine models of SSc in a transforming growth factor-ß (TGF-ß)/Smad-dependent manner. Overexpression of TRB3 stimulated canonical TGF-ß signalling and induced an activated phenotype in resting fibroblasts. In contrast, knockdown of TRB3 reduced the profibrotic effects of TGF-ß and decreased the collagen synthesis. Moreover, siRNA-mediated knockdown of TRB3 exerted potent antifibrotic effects and ameliorated bleomycin as well as constitutively active TGF-ß receptor I-induced fibrosis with reduced dermal thickening, decreased hydroxyproline content and impaired myofibroblast differentiation. CONCLUSIONS: The present study characterises TRB3 as a novel profibrotic mediator in SSc. TGF-ß induces TRB3, which in turn activates canonical TGF-ß/Smad signalling and stimulates the release of collagen, thereby inducing a positive feedback loop that may contribute to aberrant TGF-ß signalling in SSc.

Interleukin-35 is upregulated in systemic sclerosis and its serum levels are associated with early disease.

OBJECTIVES: IL-35 is a member of the IL-12 family consisting of p35/IL-12a and EBI3/IL-27b subunits. IL-35 exerts immunomodulatory activities in experimental and human autoimmune inflammatory conditions. Our aim was to assess IL-35 expression in the skin and circulation of SSc patients and to characterize its potential association with SSc-related features. METHODS: Expression of IL-35 in skin and dermal fibroblasts was quantified by quantitative PCR, immunohistochemistry and immunofluorescence. Serum levels of IL-35 (by ELISA), CRP (by turbidimetry), ANA (by immunofluorescence) and autoantibodies of the ENA complex (by immunoblot) were measured in 40 SSc patients. Serum IL-35 was determined in 40 age- and sex-matched healthy controls. RESULTS: IL-35 expression was increased in SSc skin and dermal fibroblasts in a TGF-ß-dependent manner. IL-35 induced an activated phenotype in resting fibroblasts and enhanced the release of collagen. IL-35 serum levels were increased in patients with SSc compared with healthy controls [median 83.9 (interquartile range 45.1-146.1) vs 36.2 (interquartile range 17.2-49.4) pg/ml, P < 0.0001]. Serum IL-35 was negatively correlated with disease duration (r = -0.4339, P = 0.0052). In line with this finding, serum IL-35 was increased in patients with an early SSc pattern on capillaroscopy assessment compared with those with active and late SSc patterns. CONCLUSION: The present study demonstrates overexpression of IL-35 in SSc skin, dermal fibroblasts and serum. TGF-ß induces IL-35, which in turn activates resting fibroblasts and enhances the release of collagen, thereby contributing to aberrant TGF-ß signalling in SSc. Increased serum IL-35 is associated with early, inflammatory stages of SSc.